Delivery of proteins as therapeutics or for inducing an immune response in biologically relevant amounts has been an obstacle to drug and vaccine development for decades. One solution that has proven to be a successful alternative to traditional drug delivery approaches is delivery of exogenous nucleic acid sequences for production of therapeutic factors in vivo. Gene transfer vectors ideally enter a wide variety of cell types, have the capacity to accept large nucleic acid sequences, are safe, and can be produced in quantities required for treating patients. Viral vectors have these advantageous properties and are used in a variety of protocols to treat or prevent biological disorders.
Adenoviral vectors are attractive for gene transfer applications, such as gene therapy and vaccines as a result of their ability to infect a variety of cell types with high efficiency. Adenoviral vectors containing a heterologous transgene under the control of astrong promoter are potent, achieving expression of the heterologous protein up to 20% of total cell proteins (see, e.g., Massie et al., J. Virol., 72, 2289-2296 (1998)). A high level of transgene expression, however, often is inhibitory to virus growth, such as when the transgene encodes a protein that is cytotoxic to a packaging cell. Thus, high expression of an adenovirus-encoded transgene can prevent the production of viable adenoviral vector particles from naked DNA (see, e.g., Matthews et al., J. Gen. Virol., 80 (Pt 2), 345-353 (1999)), or reduce the productivity of virus growth within packaging cells (see, e.g., Molin et al., J. Virol., 74, 9002-9009 (2000)).
To better regulate transgene expression within virus packaging cells while maintaining vector potency, gene regulation systems have been employed in the construction of adenoviral vectors. These systems typically incorporate transcriptional regulatory proteins into the adenoviral vector or in the target cell (see, e.g., Massie et al., supra, Goukassian et al., FASEB J., 15, 1877-1885 (2001), Mizuguchi et al., Biochim. Biophys. Acta, 1568, 21-29 (2001), Rubinchik et al., Mol. Ther., 4, 416-426 (2001), Molin et al., J. Virol., 72, 8358-8361 (1998), Hu et al., Cancer Res., 57, 3339-3343 (1997), Edholm et al., J. Virol., 75, 9579-9584 (2001), and U.S. Pat. No. 6,391,612). Such gene regulation systems, however, often require the use of inducer compounds (e.g., tetracycline analogs), which increases the time required to generate viable adenoviral vector particles, thereby complicating their widespread use.
Therefore, there remains a need for more efficient methods of propagating adenoviral vectors encoding transgenes whose expression inhibits viral growth in host cells. The invention provides such a method. These and other advantages of the invention, as well as additional inventive features, will be apparent from the description of the invention provided herein.